We aim to investigate the mechanisms of desensitization and long-term tolerance associated with sublingual and subcutaneous allergen immunotherapy (SLIT and SCIT, respectively). The proposed work will include investigation of allergen-specific T cells enriched and analysed freshly ex vivo using tetramers labelled with grass pollen immunodominant epitopes as well as with transcriptome analysis. Using these technologies we will primarily investigate mechanisms of SLIT, with subjects receiving SCIT or placebo acting as positive and negative control groups, respectively. This work is to be performed in the context of an Immune Tolerance Network funded randomized double-blind, double dummy, controlled clinical trial (SLIT vs. placebo, SCIT vs. placebo). Although the primary focus and endpoint of this trial relates to the effects of SLIT vs. placebo, it will provide an unparalleled opportunity to examine both forms of treatment and to correlate immunologic and clinical outcomes. Investigations will be performed both during the 'desensitisation phase'(i.e. while patients are actively receiving immunotherapy) and during the 'tolerance phase'i.e. after discontinuation of treatment. This work also has the potential to inform selection of novel biomarkers for effective SLIT and SCIT. We hypothesize that grass pollen immunotherapy (SLIT and SCIT but not placebo) will: 1. Reduce the frequencies of circulating grass pollen tetramer-reactive T cells. 2. Alter phenotypes of grass pollen tetramer-reactive Th2 cells in favor of antigen-specific Tregs and Th1 cells. 3. Induce immunologic changes that persist following immunotherapy discontinuation. 4. Modify transcriptional profiles of allergen-specific T cells, giving rise to potential new biomarkers for effective tolerization. PUBLIC HEALTH RELEVANCE (provided by applicant): Hay fever affects around 1 in 5 Americans;many respond poorly to treatments like antihistamines, but immunotherapy ('allergy shots') can be very effective. A new type of immunotherapy has been developed that is taken under the tongue as a tablet instead of by injection. We will test if this works by changing the immune system in a group of people receiving these new tablets as well as in people receiving normal shots.